1) look for your protein of interest (for example, http://www.uniprot.org/), get DNA sequence and order to to IDT or similar. insert your DNA in a plasmid of interest
-- 2) enzymes. enzymes everywhere. crispr is just a method very usefull for some organism or goals. if you just want to purify a protein from bacteria, you shouldnt worry about crispr
3) extract the dna from your transformed bacteria and run a pcr or check it though selective medium (lacZ, ampicillin...). this way, you'll know if you bacteria have incorported the dna. to check if is really your protein of interest, best way is to tun a western blot or adding a tag to your protein and purify the protein using that tag.
please, check bibliography ALWAYS
El miércoles, 23 de marzo de 2016, 6:02:21 (UTC+1), Justin S escribió:
El miércoles, 23 de marzo de 2016, 6:02:21 (UTC+1), Justin S escribió:
Hi all,I'm a total beginner, but I want to get started doing some experimental biology of my own. I'm hoping the community here can help fill in some gaps for me!1) Suppose I want to modify the DNA of some bacteria with the goal of producing some protein. How does one go about figuring out what sequence needs to be added to the bacterial genome?2) How is the sequence inserted? I'm aware of crispr, but I'm guessing there are other techniques. What's available to the layperson?3) How does one verify the desired proteins are produced?
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